The fluorescent properties of terbium (Tb3+) were used to study the calcium (Ca2+) binding sites on GH3 pituitary tumor cells. The fluorescence emission of Tb3+ was enhanced with the binding of GH3 cells, accompanied by a red shift in its excitation maximum to resemble the excitation peak of the native cell fluorescence. The Tb3+ fluorescence enhancement increased with increasing concentrations of GH3 cell protein. Scatchard plots revealed at least two classes of Tb3+ binding sites on GH3 cells. The low and high affinity binding sites have apparent dissociation constants equal to 0.56 mM and 11 microM, respectively. The high affinity Tb3+ binding was displaced by Ca2+, but the more abundant low affinity site was not sensitive to Ca2+. The data suggest that GH3 cells possess a specific Ca2+ binding receptor on their plasma membrane.